We came to the conclusion that the study of cell-mediated immune responses per se is not going to resolve the cause of MHC restrictions in T-cell killings of targets; dual recognition versus altered-self recognition. This certainly applies to cytotoxic female T-cells directed against testis-organizing H-Y antigen. The proposed physical association between H-Y antigen and beta2-microglobulin-MHC antigen dimers can only be tested by chemical means. Nevertheless, in the absence of H-Y antigen's proposed anchorage site, beta2-microglobulin (-), HLA(-) Daudi human male Burkitt lymphoma cells failed to stably maintain H-Y antigen on their plasma membrane. Instead, free H-Y antigen was found in the culture medium which enabled us to identify human H-Y antigen as a discrete protein species; polymers formed by S-S binds of the hydrophobic subunit made of 171 amino acid residues. We have already found the binding affinity between purified H-Y and bovine beta2-microglobulin to be negligible; Kd greater than 10 to the minus 2nd power M. We intend to study that between H-Y antigen and beta2m-H-2 or HLA antigen dimers in vitro.